The overwhelming necessity for effective and causal treatment of allergic diseases is well known in the art. Within the large group of allergic diseases, which are in general characterized by versatile hypersensitivity type I reactions caused by a disordered activation of the immune system and which comprise allergic rhinitis, asthma bronchiale, atopic eczema, anaphylaxis, insect venom, drug allergies, food allergies and multiple allergies, allergic airway diseases such as asthma are among the most prevalent ones to impair quality of life and life expectation. In the last decades, allergic airway diseases have dramatically increased in the northern hemisphere of industrialized countries humans and domestic animals such as horses (Kline 2007; Braun-Fähränder 2009). As of 2009, 300 million people worldwide were affected by asthma leading to approximately 250,000 deaths per year. It is estimated that asthma has a 7-10% prevalence worldwide with a great disparity in prevalence worldwide across the world (as high as a 20 to 60-fold difference). A trend toward more developed and westernized countries having higher rates of asthma was observed.
Depending on the severity, allergic airway diseases such as asthma are associated with inflammation and airway obstruction. However, even if some symptoms might be comparable, the underlying principles causing the disease are different from chronic obstructive pulmonary disease (COPD). Nevertheless, a medication to both control allergy and to reduce inflammation would be of great general advantage.
In present medication of asthma one can be distinguished between fast acting and long term control. Short acting beta 2-adrenoceptor agonists (SABA), such as salbutamol are the gold standard in treatment of asthma symptoms. Anticholinergic medications such as ipratropium bromide provide addition benefit when used in combination with SABA in those with moderate or severe symptoms. Anticholinergic bronchodilators are an alternative if a person cannot tolerate a SABA.
For long term control, glucocorticoids are considered the most effective treatment available. Inhaled forms are usually used except in the case of severe persistent disease, in which oral steroids appear appropriate. Inhaled formulations may be used once or twice daily, depending on the severity of symptoms. Long acting beta-adrenoceptor agonists (LABA) have at least a 12-hour effect. They are however not to be used without a steroid due to an increased risk of severe symptoms and are thus challenged for additive value. Leukotriene antagonists such as montelukast or zafirlukast are an alternative to inhaled glucocorticoids, but are second line at present. Mast cell stabilizers such as cromolyn sodium are another but less potent alternative to glucocorticoids. Anti-immunoglobulin E (IgE) monoclonal antibodies constitute a relatively new but not yet broadly established potentially causal from of antiallergic medication.
Despite such advanced knowledge and various available mediation regimens, the increasing frequency of asthma is alarming. Rates of asthma have increased significantly between the 1960s and 2008 with 9% of US children suffering from asthma in 2001, compared with just 3.6% in 1980. The World Health Organization (WHO) reports that today 10% of the Swiss population suffers from asthma today compared with just 2% some 25-30 years ago. Thus, the establishment of alternative causal medication strategies is highly desirable and required.
Allergic airway diseases are not restricted to humans. Like human asthma, recurrent airway obstruction (RAO) in horses is considered a multifactor allergic airway hypersensitivity reaction elicited by environmental exposure to potential allergens (Robinson 2001) and heritable components (Gerber et al. 2009). Equine RAO has become one of the most common airway diseases (Fey 2006). Housing of horses in stables with permanent exposure to potentially allergenic organic and inorganic particles was reported to be a major trigger factor (Schmallenbach et al. 1998; Robinson 2001; Millerick-May 2009). Keeping horses on pasture leads to improved clinical signs, however, complete avoidance of allergens is not always possible (Robinson 2001). The permanent inhalation of various antigens from moldy hay, mite dust as well as endotoxin, β-glucan and other organic and inorganic particles causes airway neutrophilia and inflammation with a mixed Th1/Th2 immune response (Horohov et al. 2005; Cordeau et al. 2004). Although the clinical signs of RAO were well defined (Robinson 2001), immunological mechanisms are still controversy discussed. However, a predominant Th2 allergic response was recently presumed (Horohov et al. 2009). In several studies, signs of predominant Th2 response such as high IL-4, IL-5 in bronchoalveolar lavage fluid (BALF) of RAO horses exposed to antigens were reported (Lavoie et al. 2001; Cordeau et al. 2004). Due to persistent chronic inflammatory reaction in small airways of the affected horses, Th1 participation was also confirmed (Ainsworth et al. 2003).
Unmethylated Cytosin-Phosphate-Guanin-Oligodeoxynucleotides (CpG-ODN) were described as effective immune stimulating agents to cause a Th2/Th1 immune shift (Kline 2007). This shift further promoted an immunoglobuline isotype switch from IgE to IgG2 (Bohle 2002). The prevalence of IgE is a matter of ongoing debate (Halliwell et al. 1993; Marti 2009). In addition, a shift from pro-allergy mediating IL-4, IL-5 and IL-13 Th2 cytokines towards pro-inflammatory IFN-γ and IL-12 Th1 cytokines was discussed. Anti-inflammatory and antiallergic properties of Th2 cytokine IL-10 turned out to be of interest. Particularly, IL-10 producing T regulatory cells (Tregs) and its balance towards Th2 cells seems to play an important role in immune homeostasis (Akdis et al. 2004; Lloyd and Hawrylowicz 2009). Thus, one problem to be solved by the present invention is the clarification whether alternatively or in addition to the present conventional symptomatic therapy the possibility to modulate cytokine level in order to avoid development of an allergic hypersensitivity constitutes a promising option for use in the treatment of allergic and/or inflammatory lower airway diseases.
However, so far no decisive IL-10 regulating nanoparticulate composition comprising CpG-ODN has been proposed for use in therapy in allergic and inflammatory airway diseases, especially not my means of inhalation. Hence, there is a need for immunomodulating nucleic acids in a nanoparticulate composition for use in the treatment of allergic and/or inflammatory airway diseases such as asthma and RAO.
Three distinctive CpG classes (A-, B- and C-class) with varying immunologic effects were previously investigated (Krieg 2002). The key pattern-recognition receptor for these “danger signals” is the Toll-Like receptor 9 (TLR-9) which is located in endosomes (Krieg 2002). Recently, equine TLR-9 was detected in monocytes, airway epithelial cells, capillary endothelium in the lungs and pulmonary intravascular macrophages (Schneeberger et al. 2009). Furthermore, species-specific immune stimulation depending on CpG motifs has been determined (Hartmann and Krieg 2000; Rankin et al. 2001). US2004/0132682 claims an interesting method of treating inflammatory lung disease with suppressors of CpG-ODNs. However, these inflammatory lung diseases were explicitly not of allergic origin. The person of ordinary skill appreciates that the immunologic processes underlying allergic and non-allergic inflammatory inflammation of the lung are distinctive, e.g. concerning the involvement of neutrophils versus eosinophils.
Nevertheless, a promising immunotherapeutic strategy against allergic conditions of the upper airways including the nose as well as of the eyes already entering human clinical phase IIa studies involved CpG-ODN (Senti et al. 2009) as adjuvant, i.e. a synthetic analog of natural microbial CpG-ODN (Krieg 2002). This TLR9 agonist demonstrated some efficiency in treatment of allergic diseases if combined as an adjuvant with specific allergens due to CpG-DNA's potential to cause a Th2/Th1 shift (Vollmer and Krieg 2009, Kline 2007, Krieg 2002) which was associated with a downregulation of proallergic Th2 cytokines (IL-4, IL-5, IL-13) and an upregulation of antiallergic Th1 cytokines (IFN γ, IL-12) and an immunoglobulin isotype switch from allergy mediating IgE to IgG (Bohle 2002). U.S. Pat. No. 6,429,199 claims such CpGs as adjuvants in immunotherapy. The role of CpGs as adjuvants in order to stimulate the immune system is further supported by U.S. Pat. No. 7,488,490. However, despite for treatment of non-allergic lung diseases (US2004/0132682) such prior art rather teaches away from the employment of CpG-ODNs in monotherapy, especially for inhalation therapy, as inhalation of CpG-ODNs without immunotherapy did not result in a reduction of allergic clinical symptoms in a clinical trial (Fonseca et al 2009). Moreover, prior art is silent on an immunmodulating action based on IL-10 stimulation and especially, nanoparticles in no form are disclosed as eligible for use in related immunomodulating therapy, neither in general nor specifically for inhalative administration.
Pulmonary delivery as a non-invasive route of drug administration still constitutes a vivid research field in the treatment and diagnosis of respiratory and non-respiratory diseases (Smola et al. 2008). However, nanoparticulate delivery is still a partly unexplored field.
As an example for protein-based nanoparticles, gelatin nanoparticles (GNPs) from gelatin type A obtained from porcine skin were found to be immunologically inert and the use of GNPs showed amplified CpG-related TLR-9 activation for use as adjuvant in anticancer immunotherapy (Zwiorek et al. 2008). However, none such composition was ever evaluated in reducing the immune system's activity, for example for effective use in the treatment of allergic and/or inflammatory diseases. In detail, one problem to be solved is to identify a composition comprising the optimal immunomodulating agent to trigger specific immunomodulating effect such as Th2/Th1 shift and IL-10 stimulation and to reduce, for example, inflammation in clinical allergic symptoms known to the skilled person
Compressor or jet nebulizers apparently are not suitable for nebulizing compositions comprising fragile nucleic acid such as CpG-ODNs even if jet nebulized protein-based nanoparticles such as from gelatin gave promising results such as enabling targeted lung cancer delivery of chemotherapeutic agent cisplatin (Tseng et al. 2007; Tseng et al. 2009). However, these compositions were much more stable than any nucleic acid comprising nanoparticulate composition. Tseng determined the median droplet size and lung deposition was proved in vivo, but no further characterization of the data of the nebulization process itself was given. US2005/019270 describes a method to nebulize spray-dried GNPs leaving the skilled person in doubt whether such methods destroys sensitive payload such as CpG-ODN. Against this technical background and the fact that the frequency of allergic and inflammatory airway diseases is still increasing, the problem is that there remains a tremendous need for new, long-lasting and efficient composition for use in causal prevention and/or treatment of allergic and inflammatory airway diseases.